Abstract (eng)
Plasmalogens are special ether lipids that differ in having a vinyl ether bond at their sn1 position and in being abundant in membrane rafts. Plasmalogen deficiency leads to the rare
inherited disorder rhizomelic chondrodysplasia punctata (RCDP) that, in some cases, is also accompanied by heart defects. In non‐RCDP patients, such cardiac defects have been associated with a downregulation and/or mislocalization of Connexin‐43 (Cx43). This protein has been shown to be reduced in fibroblasts of ether lipid‐deficient mice.
The aim of the present thesis was to detect a possible molecular link between Cx43 and
ether lipids in RCDP cells and hearts of mice deficient in ether lipids.
We were able to distinguish the non‐phosphorylated and phosphorylated isoforms of Cx43 in our system. The non‐phosphorylated isoform represents newly synthesized intracellular protein, while phosphorylated Cx43 isoforms are at the cell surface or in gap junctions. Our results demonstrate a confluency‐dependent expression of Cx43. This protein was upregulated in human primary control fibroblasts upon confluency. However, this induction was not observed in ether lipid‐deficient fibroblasts. These cells showed a significant reduction in Cx43 protein levels compared to control. Interestingly, Cx43 was particularly reduced in its phospho‐isoforms. Furthermore, immunofluorescence staining revealed Cx43 to be localized exclusively at sites of cell‐cell contact in control cells, resulting in a continuous signal. Human primary fibroblasts lacking ether lipids did not exhibit this pattern. We also examined localization of Cx43 in rafts by isolating detergent‐resistant membranes from both primary cell lines and detected most of the Cx43 phospho‐isoforms to be in raft fractions of control cells, whereas in ether lipid‐deficient cells they were merely present. Due to the many reported RCDP cases associated with heart anomalies, mouse heart tissues were examined. Hearts of mice lacking a crucial biosynthetic enzyme for ether
lipid biosynthesis also showed significantly lower amounts of Cx43.
Taken together, this study points towards a possible involvement of Cx43 in the pathologic course of RCDP. It is also tempting to speculate that Cx43 might be involved in the molecular mechanisms underlying underlying the heart defects reported in RCDP patients. Thus, these data provide sufficient information to justify future detailed investigations.